Novel mechanisms of glomerular injury in primary membranous nephropathy Primary membranous nephropathy (MN) is the most common cause of nephrotic syndrome in adults. Injury is initiated by the deposition of circulating auto-antibodies against podocyte antigens in the subepithelial space of the basement membrane, leading to complement activation and formation of the C5b-9 membrane attack complex (MAC). MAC-mediated sublytic injury is thought to cause podocyte damage and glomerular disruption. However, a randomized controlled trial showed no benefit of anti-C5 antibody in disease remission in patients with MN, suggesting that other MAC-independent mechanisms apply. Studies on MN pathophysiology have been limited by the lack of reliable in vitro systems. We generated a new glomerulus-on-a-chip platform that we will use as a tool to assess alternative mechanisms of injury in MN. Preliminary data suggest that complement activation leads to C3a/C3aR signaling in podocytes, which induces loss of glomerular permselectivity due to SNAIL-mediated signaling. Moreover, challenging current assumption in MN, we have found that MN-serum leads to C3a/C3aR signaling also in glomerular endothelial cells and induces overexpression of MMP9, thus suggesting a role of this signaling in regulating glomerular basement membrane-cell interaction. These effects are prevented by a C3aR antagonist, suggesting the direct involvement of C3a in podocyte and GEC damage and in glomerular basement membrane (GBM) degradation. Therefore, based on our preliminary data we hypothesize that C3a/C3aR signaling not only plays a critical role in podocyte damage but exert its effects also on GEC, leading to disruption of glomerular filtration barrier and loss of permselectivity. To test our hypothesis, we will investigate the effects of C3a signaling, its mechanism(s) of action and its role in GBM degradation both in vitro and in vivo. If confirmed, our findings will not only provide novel evidence that multiple injury mechanisms are in play in PMN but will also inform us about potential therapeutic targets that could prove useful for the development of new treatments for MN and other proteinuric diseases.